Beta-Amino acids are rapidly growing in importance as a key class of pharmaceutical intermediates, with applications in a number of current and future drugs. A cost-effective process for the production of optically pure beta-amino acids will be developed in this project to accommodate the commercial needs for beta-amino acids. The method involves two enzymatic reactions starting from conveniently prepared racemic substituted dihydrouracils. The 5,6-dihydrouracil, which may be substituted at either the 5 or 6 positions with various functional groups, is contacted with a dihydrouracilase enzyme to catalyze a stereoselective hydrolysis reaction, producing an enantiomerically-enriched chiral N-carbamoyl-beta-amino acid and an enantiomerically enriched, unreacted chiral 5,6-dihydrouracil. The chiral N-carbamoyl-(-amino acid was then decarbamoylated enzymatically to produce an optically pure (-amino acid. The opposite enantiomer of the beta-amino acid can be produced in pure form by recovering and hydrolyzing the unreacted chiral 5,6- dihydrouracil using the enzymes with opposite stereoselectivity. An important feature of this method is that the two enzyme reaction can be carried out in one pot and it is not necessary to isolate the intermediate N-carbamoyl-beta-amino acids. This proof of concept has been demonstrated in Phase 1 using 5,6-dihydrouracil and 5-methyl-5,6-dihydrouracil. In Phase II, the top priority will be searching for carbamoylase enzymes with a broader substrate range, particularly including aryl-substituted N-carbamoyl-(-amino acids using directed evolution technology. Dihydrouracilases with opposite enantioselectivity (i.e. enantioselectively hydrolyzing the R-enantiomer of dihydrouracils) will also be found and produced. The cost-effective one-pot process to convert racemic substituted dihydrouracils to the corresponding enantiomerically pure beta-amino acids via the coupled dihydrouracilase/carbamoylase two-enzyme system in the non-immobilized and immobilized forms will be developed further using the enzymes acquired in Phase 2. Finally, the production of enantiomerically-pure beta-amino acids will be demonstrated on the 100 gram scale for important commercial targets.